Respiratory syncytial virus (RSV) is the major cause of severe lower respiratory tract infections in children and infants, affecting an estimated 64 million people and causing 160,000 deaths worldwide per year (WHO). Each year in the United States, ~100,000 infants and ~170,000 adults are hospitalized due to RSV infection. High-risk adults (elderly and patients with chronic heart or lung disease) experience the highest RSV-attributed mortality in the US (~14,000 deaths/year), similar to the rates of seasonal influenza. The annual cost of RSV on the US healthcare system is ~$2 billion. Currently, no vaccines or disease-specific therapeutics are available to combat RSV, and treatment is mostly limited to supportive care. For the most acute cases, ribavirin, a broad-range antiviral with questionable efficacy and significant safety issues can be used, though it is not indicated for adults. Only one preventative is available, Synagis (Palivizumab), a monoclonal antibody that inhibits RSV infection, but its use is limited to high-risk infants. Synagis is very expensive (~$5000 for a course of therapy) and reduces RSV-related hospitalizations of high-risk infants by only 55%. There is an urgent need for new anti-RSV prophylactics and therapies that can be applied to a broad patient population. In this project we will apply an innovative strategy to identify novel, protease-resistant D-peptide drug candidates for the critically underserved RSV patient population. Our drug discovery platform employs an enantiomeric screening technology (mirror-image phage display) coupled with protein design, to identify D- peptides that stop the virus as it attempts to enter a cell. We have successfully validated this platform technology by identifying a promising anti-HIV preclinical candidate, which is the most specific and potent D- peptide inhibitor known. Our anti-HIV D-peptide targets a conserved pocket found on a region of the HIV envelope protein, the N-trimer, which is transiently exposed during viral entry. It inhibits all maor circulating HIV-1 strains and, by design, possesses an extremely high barrier to resistance. RSV uses a highly similar mechanism of viral entry, and an analogous vulnerable pocket on the N-trimer has been identified on the RSV viral surface. This pocket will be the target for our discovery efforts. In this two-year Phase I SBIR, we propose to identify and structurally characterize 1st generation D-peptide RSV entry inhibitors. In Phase II we will rapidly apply all of the optimization techniques used in our HIV discovery efforts, which improved the potency of our HIV-specific drug by over six orders of magnitude and gave it an extraordinary resistance barrier. Once we have fully optimized the potency, specificity and resistance profile of our D-peptide RSV entry inhibitor, we will explore its utility as an anti-RSV therapeutic and preventative.